Mitotic control of RUNX2 phosphorylation by both CDK1/cyclin B kinase and PP1/PP2A phosphatase in osteoblastic cells

Arun Rajgopal, Daniel W. Young, Khawaja A. Mujeeb, Janet L. Stein, Jane B. Lian, Andre J van Wijnen, Gary S. Stein

Research output: Contribution to journalArticle

45 Citations (Scopus)

Abstract

Skeletal development and osteoblast maturation require the phenotype promoting activity of the transcription factor RUNX2, which controls both cell growth and differentiation in osteoblasts. We have recently shown that in actively proliferating cells RUNX2 regulates the expression of specific target genes as cells enter and exit mitosis. In this study, we addressed whether post-translational modifications of RUNX2 control its activity during mitotic exit. Western blot analysis of proteins from osteoblastic Saos-2 cells released from mitotic inhibition into early G1 show a phosphatase-sensitive shift in the mobility of RUNX2 in SDS gels. The slowly migrating hyper-phosphorylated form of RUNX2 is immunoreactive with a CDK related phospho-antibody (MPM2) only in mitotic cells and is converted into a faster migrating hypo-phosphorylated RUNX2 when cells complete mitosis. This conversion is inhibited by okadaic acid, an inhibitor of protein phosphatases 1 and 2 (PP1 and PP2A), but not by deltamethrin which blocks PP2B phosphatase. Mitotic phosphorylation of RUNX2 is sensitive to the CDK inhibitors roscovitine and olomoucine. Furthermore, RUNX2 can directly interact with CDK1 and is phosphorylated in vitro by the CDK1/cyclin B kinase complex. Hence, RUNX2 is hyper-phosphorylated by CDK1/cyclin B during mitosis, and dynamically converted into a hypo-phosphorylated form by PP1/PP2A-dependent dephosphorylation after mitosis to support the post-mitotic regulation of RUNX2 target genes.

Original languageEnglish (US)
Pages (from-to)1509-1517
Number of pages9
JournalJournal of Cellular Biochemistry
Volume100
Issue number6
DOIs
StatePublished - Apr 15 2007
Externally publishedYes

Fingerprint

Cyclin B
Phosphorylation
Osteoblasts
Phosphoric Monoester Hydrolases
Phosphotransferases
Mitosis
Genes
Okadaic Acid
Cell growth
Transcription Factors
Gels
Antibodies
Post Translational Protein Processing
Proteins
Cell Differentiation
Western Blotting
Phenotype
Growth
olomoucine
decamethrin

Keywords

  • CDK1
  • Cyclin B
  • Mitosis
  • Okadaic acid
  • Osteoblast
  • Osteosarcoma
  • Roscovitine
  • RUNX2

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

Cite this

Rajgopal, A., Young, D. W., Mujeeb, K. A., Stein, J. L., Lian, J. B., van Wijnen, A. J., & Stein, G. S. (2007). Mitotic control of RUNX2 phosphorylation by both CDK1/cyclin B kinase and PP1/PP2A phosphatase in osteoblastic cells. Journal of Cellular Biochemistry, 100(6), 1509-1517. https://doi.org/10.1002/jcb.21137

Mitotic control of RUNX2 phosphorylation by both CDK1/cyclin B kinase and PP1/PP2A phosphatase in osteoblastic cells. / Rajgopal, Arun; Young, Daniel W.; Mujeeb, Khawaja A.; Stein, Janet L.; Lian, Jane B.; van Wijnen, Andre J; Stein, Gary S.

In: Journal of Cellular Biochemistry, Vol. 100, No. 6, 15.04.2007, p. 1509-1517.

Research output: Contribution to journalArticle

Rajgopal, A, Young, DW, Mujeeb, KA, Stein, JL, Lian, JB, van Wijnen, AJ & Stein, GS 2007, 'Mitotic control of RUNX2 phosphorylation by both CDK1/cyclin B kinase and PP1/PP2A phosphatase in osteoblastic cells', Journal of Cellular Biochemistry, vol. 100, no. 6, pp. 1509-1517. https://doi.org/10.1002/jcb.21137
Rajgopal, Arun ; Young, Daniel W. ; Mujeeb, Khawaja A. ; Stein, Janet L. ; Lian, Jane B. ; van Wijnen, Andre J ; Stein, Gary S. / Mitotic control of RUNX2 phosphorylation by both CDK1/cyclin B kinase and PP1/PP2A phosphatase in osteoblastic cells. In: Journal of Cellular Biochemistry. 2007 ; Vol. 100, No. 6. pp. 1509-1517.
@article{51c82ffcd114482d88a521e51a027b95,
title = "Mitotic control of RUNX2 phosphorylation by both CDK1/cyclin B kinase and PP1/PP2A phosphatase in osteoblastic cells",
abstract = "Skeletal development and osteoblast maturation require the phenotype promoting activity of the transcription factor RUNX2, which controls both cell growth and differentiation in osteoblasts. We have recently shown that in actively proliferating cells RUNX2 regulates the expression of specific target genes as cells enter and exit mitosis. In this study, we addressed whether post-translational modifications of RUNX2 control its activity during mitotic exit. Western blot analysis of proteins from osteoblastic Saos-2 cells released from mitotic inhibition into early G1 show a phosphatase-sensitive shift in the mobility of RUNX2 in SDS gels. The slowly migrating hyper-phosphorylated form of RUNX2 is immunoreactive with a CDK related phospho-antibody (MPM2) only in mitotic cells and is converted into a faster migrating hypo-phosphorylated RUNX2 when cells complete mitosis. This conversion is inhibited by okadaic acid, an inhibitor of protein phosphatases 1 and 2 (PP1 and PP2A), but not by deltamethrin which blocks PP2B phosphatase. Mitotic phosphorylation of RUNX2 is sensitive to the CDK inhibitors roscovitine and olomoucine. Furthermore, RUNX2 can directly interact with CDK1 and is phosphorylated in vitro by the CDK1/cyclin B kinase complex. Hence, RUNX2 is hyper-phosphorylated by CDK1/cyclin B during mitosis, and dynamically converted into a hypo-phosphorylated form by PP1/PP2A-dependent dephosphorylation after mitosis to support the post-mitotic regulation of RUNX2 target genes.",
keywords = "CDK1, Cyclin B, Mitosis, Okadaic acid, Osteoblast, Osteosarcoma, Roscovitine, RUNX2",
author = "Arun Rajgopal and Young, {Daniel W.} and Mujeeb, {Khawaja A.} and Stein, {Janet L.} and Lian, {Jane B.} and {van Wijnen}, {Andre J} and Stein, {Gary S.}",
year = "2007",
month = "4",
day = "15",
doi = "10.1002/jcb.21137",
language = "English (US)",
volume = "100",
pages = "1509--1517",
journal = "Journal of Cellular Biochemistry",
issn = "0730-2312",
publisher = "Wiley-Liss Inc.",
number = "6",

}

TY - JOUR

T1 - Mitotic control of RUNX2 phosphorylation by both CDK1/cyclin B kinase and PP1/PP2A phosphatase in osteoblastic cells

AU - Rajgopal, Arun

AU - Young, Daniel W.

AU - Mujeeb, Khawaja A.

AU - Stein, Janet L.

AU - Lian, Jane B.

AU - van Wijnen, Andre J

AU - Stein, Gary S.

PY - 2007/4/15

Y1 - 2007/4/15

N2 - Skeletal development and osteoblast maturation require the phenotype promoting activity of the transcription factor RUNX2, which controls both cell growth and differentiation in osteoblasts. We have recently shown that in actively proliferating cells RUNX2 regulates the expression of specific target genes as cells enter and exit mitosis. In this study, we addressed whether post-translational modifications of RUNX2 control its activity during mitotic exit. Western blot analysis of proteins from osteoblastic Saos-2 cells released from mitotic inhibition into early G1 show a phosphatase-sensitive shift in the mobility of RUNX2 in SDS gels. The slowly migrating hyper-phosphorylated form of RUNX2 is immunoreactive with a CDK related phospho-antibody (MPM2) only in mitotic cells and is converted into a faster migrating hypo-phosphorylated RUNX2 when cells complete mitosis. This conversion is inhibited by okadaic acid, an inhibitor of protein phosphatases 1 and 2 (PP1 and PP2A), but not by deltamethrin which blocks PP2B phosphatase. Mitotic phosphorylation of RUNX2 is sensitive to the CDK inhibitors roscovitine and olomoucine. Furthermore, RUNX2 can directly interact with CDK1 and is phosphorylated in vitro by the CDK1/cyclin B kinase complex. Hence, RUNX2 is hyper-phosphorylated by CDK1/cyclin B during mitosis, and dynamically converted into a hypo-phosphorylated form by PP1/PP2A-dependent dephosphorylation after mitosis to support the post-mitotic regulation of RUNX2 target genes.

AB - Skeletal development and osteoblast maturation require the phenotype promoting activity of the transcription factor RUNX2, which controls both cell growth and differentiation in osteoblasts. We have recently shown that in actively proliferating cells RUNX2 regulates the expression of specific target genes as cells enter and exit mitosis. In this study, we addressed whether post-translational modifications of RUNX2 control its activity during mitotic exit. Western blot analysis of proteins from osteoblastic Saos-2 cells released from mitotic inhibition into early G1 show a phosphatase-sensitive shift in the mobility of RUNX2 in SDS gels. The slowly migrating hyper-phosphorylated form of RUNX2 is immunoreactive with a CDK related phospho-antibody (MPM2) only in mitotic cells and is converted into a faster migrating hypo-phosphorylated RUNX2 when cells complete mitosis. This conversion is inhibited by okadaic acid, an inhibitor of protein phosphatases 1 and 2 (PP1 and PP2A), but not by deltamethrin which blocks PP2B phosphatase. Mitotic phosphorylation of RUNX2 is sensitive to the CDK inhibitors roscovitine and olomoucine. Furthermore, RUNX2 can directly interact with CDK1 and is phosphorylated in vitro by the CDK1/cyclin B kinase complex. Hence, RUNX2 is hyper-phosphorylated by CDK1/cyclin B during mitosis, and dynamically converted into a hypo-phosphorylated form by PP1/PP2A-dependent dephosphorylation after mitosis to support the post-mitotic regulation of RUNX2 target genes.

KW - CDK1

KW - Cyclin B

KW - Mitosis

KW - Okadaic acid

KW - Osteoblast

KW - Osteosarcoma

KW - Roscovitine

KW - RUNX2

UR - http://www.scopus.com/inward/record.url?scp=34047117981&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=34047117981&partnerID=8YFLogxK

U2 - 10.1002/jcb.21137

DO - 10.1002/jcb.21137

M3 - Article

C2 - 17171635

AN - SCOPUS:34047117981

VL - 100

SP - 1509

EP - 1517

JO - Journal of Cellular Biochemistry

JF - Journal of Cellular Biochemistry

SN - 0730-2312

IS - 6

ER -